Gel electrophoresis is a well known method for separation and analysis of proteins, nucleic acids and other charged species. For purposes of this application by electrophoresis is meant the category of separation methods which utilizes an applied voltage to cause migration of charged species in an anti-convection medium, including but not limited to electrophoresis, immunoelectrophoresis, SDS (sodium dodecylsulfate) electrophoresis, isoelectric focusing, affinoelectrophoresis, 2-dimensional electrophoresis and the like.
Many electrophoretic procedures are advantageously performed in thin layers of gel on glass plates. For convenience the gel layer is pre-cast into a rectangular shape of uniform thickness affixed to a glass plate substrate of substantially the same size. The pre-cast gel covered glass surface, i.e. gel slab, frequently has a series of uniform indentations or sample wells aligned in a row at one end of the plate where samples of the substances to be electrophoresed are deposited. Uniformity in the thickness of the gel layer except where there are sample wells is an important characteristic of the gel slabs.
The use of polyacrylamide as the gel is particularly advantageous for many procedures and a variety of methods to produce such polyacrylamide gel plates have been developed. Typically, a monomer solution or other gel forming liquid is first brought in contact with its glass substrate and then polymerized or allowed to set into a relatively solid, non-flowable state. The inhibition of the polymerization of acrylamide monomer solutions by oxygen is well known. As such, acrylamide solutions are typically degassed under vacuum to remove any dissolved oxygen and then polymerized in a substantially sealed apparatus.
Various methods of casting gel slabs are known. Where the gel-forming liquid is other than polyacrylamide it is known to position a glass plate between parallel spacer rails having raised edges, the height of the edge corresponding to the thickness of the gel layer desired, to place gel-forming liquid in the center of the plate, and to smear the liquid across the entire plate using a straight edge. Such a method is not useful for casting polyacrylamide gel slabs since the acrylamide monomer will not polymerize in an apparatus wherein the liquid layer is openfaced and exposed to air. As such, separate equipment would be required for preparing polyacrylamide gel slabs and a typical laboratory which requires all varieties of gel slabs would experience duplicative equipment costs.
Whenever polyacrylamide is the gel-forming liquid, prior art procedures have required that the monomer solution be introduced into a space between plates sealed by a gasket around the edges, except for the edge into which the monomer is introduced. Typically two glass plates are separated by a rubber gasket interjected between the four edges and held together and sealed by a series of adjacent clamps. The thin space thus formed is then placed in a vertical position and monomer is introduced at one corner where the gasket is displaced. After the space is filled the remaining portion of the gasket is put in place and clamped. The monomer solution is then polymerized in place by procedures known in the art, e.g. by exposure to polymerizing light, heat or the like, depending upon the cross-linking agent and/or initator present in the solution.
Known methods of casting polyacrylamide gel slabs require cumbersome apparatus for constructing sealed spaces. Moreover, a separate sealed space must be formed for each such slab to be cast. Where molds are provided which hold more than one set of plates in an upright position each set of plates must be of equal dimensions. Where only a single sample is to be tested and thus only a narrow portion of the full sized plate is to be utilized, the experimenter has only the options of under-utilizing the plate formed or cutting it into appropriately sized strips. A further difficulty with the known procedures of gel slab preparation is that non-uniform gel layer thickness can result from uneven pressure on the plates, such as from unequal tightening of various clamps.
Accordingly there is a need for a simplified and less cumbersome method and apparatus for casting gel slabs which is applicable to the various gel-forming liquids used in electrophoretic procedure. Additionally there is a need for a method and apparatus which is easy and inexpensive and by which multiple gel slabs of varying dimensions can be simultaneously or sequentially cast.